Our objectives are to determine the genetic basis for virulence and adaptation to cell culture of HAV and to use this information to develop a strain of HAV suitable for use as an attenuated vaccine. Determinants of cell culture adaptation and of virus virulence are being identified by studying chimeric viruses encoded by infectious cDNA clones. Virulence for primates maps to two separate genes that encode nonstructural proteins. Incorporation of one virulence gene from a simian virus into the human strain of attenuated virus has produced a candidate live attenuated vaccine that is being tested in chimpanzees. A second recombinant strain that appears to have superior growth properties is being evaluated also. Since an oral attenuated vaccine would be the most efficient to administer, we are starting to define the parameters required for successful oral vaccination. We will also evaluate new routes of inoculation of infectious viral RNA. We have developed new cell lines that permit us to culture wild-type virus. The use of calf serum during vaccine production has become a health issue. We have been selecting new virus mutants and cell lines in an effort to find a virus-cell combination that will produce large amounts of virus (necessary for cost effectiveness) in the absence of serum. We have isolated chimpanzee monoclonal antibodies that neutralize HAV and will test them to determine if they can be used for passive immunoprophylaxis in humans. We have shown that an ELISA to the 3C proteinase of HAV can be used to identify infection during vaccine trials